SPECIFIC DIGOXIGENIN-LABELLED RIBOPROBES FOR DETECTION OF CITRUS PSOROSIS VIRUS AND CITRUS VARIEGATION VIRUS BY MOLECULAR HYBRIDIZATION

摘要

A dot-blot hybridization protocol using digoxigenin-labelled riboprobes was finalized for the detection of Citrus psorosis virus (CPsV) and Citrus variegation virus (CVV). Both viruses were readily identified in different organs of screenhouse-grown and, throughout a 9-month period, in-field-grown citrus plants. With CPsV, strong hybridization signals were obtained by dot blot hybridization from flowers (ovaries) and young leaves and, with CVV, from young leaves and shoots. In tissues prints, CPsV was satisfactorily detected from ovaries and CVV from petioles, ovaries, Young leaves and tender shoots. The sensitivity threshold of the assay was determined to be 75 pg of in vitro transcripts for both CPsV and CVV, allowing detection of both viruses even when their titres decreased in plant tissues and ELISA tests failed.